We have identified a new gene which when mutated leads to a form of progressive kidney disease characterized by proteinuria, reduced glomerular filtration rate, and a histologic pattern of injury characterized by focal and segmental glomerulosclerosis (FSGS). This gene, INF2 (for Inverted Formin 2), is a member of the diaphanous subgroup of the formin family. INF2, like other members of the diaphanous formin subfamily, functions to accelerate actin filament polymerization. Unlike other members, INF2 can also accelerate actin depolymerization. Members of this subfamily of formins are able to autoinhibit their activity by an intramolecular interaction between two domains, the N-terminal DID (diaphanous inhibitory domain) and the C-terminal DAD (diaphanous autoregulatory domain). We have found point mutations in the DID of INF2 in eleven unrelated families with autosomal dominant FSGS. These mutations segregate with disease, are absent from controls, and alter highly conserved amino acids. The mode of inheritance, the absence of clear loss-of-function alleles, and the localization of all of the mutations to the DID, suggest a gain-of-function effect. In this application, we propose experiments to help define the mechanism by which INF2 mutations lead to human disease and to understand the role of INF2 in kidney function. In Aim 1, we propose to explore the effects of INF2 mutations on its biochemical functions. We will examine the hypothesis that FSGS- associated mutations in INF2 disrupt intramolecular interactions leading to functional changes in INF2. We will test the effects of FSGS-causing mutations on INF2 intramolecular interactions (DID with DAD), on the ability of INF2 to mediate actin filament polymerization and depolymerization, and the interactions of INF2 with actin filaments and microtubules. In Aim 2, we will define the effects of INF2 mutations on cellular functions. We will examine the hypothesis that INF2 mutations, by disinhibiting INF2 activity, alter actin- based cell function. We will study the role of INF2 in cells and the effect of INF2 mutations on actin-based structures, on actin dynamics, on the dynamics of the endoplasmic reticulum, and on interacting proteins. In Aim 3, we will develop and analyze two new INF2 mutant mouse models. We will use these models to test the role of INF2 in the podocyte and observe the effects of an INF2 point mutation in an in vivo model. We will engineer a mouse model with a disease-associated INF2 point mutation and also develop a podocyte-specific INF2 deficient mouse. We will examine the effect of INF2 mutation and deficiency on glomerular function in vivo. PUBLIC HEALTH RELEVANCE: We have identified a new focal segmental glomerulosclerosis (FSGS) gene. When mutated, this gene, INF2, causes kidney disease in humans. Better understanding how defects in this gene cause human disease will have significant and direct implications for understanding, and ultimately, treating, common forms of renal failure and renal failure progression.